Establishment and Application of Dual TaqMan Fluorescence Quantitative Polymerase Chain Reaction (PCR) for the Detection of Porcine Circovirus Types 2 and 3

Cao, Fuliang; Luan, Xiaoning; Wang, Wenwen; Li, Yuyan; Guo, Longzong; Wang, Yinxin; Li, Jianliang

doi:10.21203/rs.3.rs-319924/v1

Cited by 2 publications

(2 citation statements)

References 27 publications

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“…In comparison, the detection limit for ddPCR was two copies/μL for PCV2 and 1 copy/μL for PCV3. The detection limits of another experimental method for PCV2 and PCV3 were ten and one copies/μL ( Cao et al, 2021 ). Therefore, the present method provided favorable anti-interference, specificity, and repeatability and is applicable to clinical or normal diagnosis and detection, early warning, and surveillance of PCV2 and PCV3.…”

Section: Discussionmentioning

confidence: 99%

Pathogenic ecological characteristics of PCV2 in large-scale pig farms in China affected by African swine fever in the surroundings from 2018 to 2021

Liu

Tong

et al. 2023

Front. Microbiol.

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Porcine circovirus type 2 (PCV2) has been identified as the causal agent of postweaning multisystemic wasting syndrome (PMWS), an economically important multifactorial disease of the swine industry worldwide. This research designed a dual nested polymerase chain reaction (PCR) detection method to simultaneously monitor porcine circovirus type 2 (PCV2) and PCV3. The limit of detection (LoD) of sensitivity for PCV2 and PCV3 was ten copies/mL for both viruses. There was no cross-reaction with any other porcine pathogens tested and no non-specific amplification. The coincidence and repetition rates were both 100%. Through the systematic and clinical sampling, 15,130 samples collected from 30 large-scale pig farms in eight provinces in China (including Hubei, Hunan, Henan, Jiangxi, Shanxi, Guangdong, Hainan, and Heilongjiang) were subjected to early warning surveillance and/or clinical diagnosis. These results revealed that the overall positive rates of PCV3 and PCV2 were 0 and 28.29%, respectively, with the lowest level recorded in Jiangxi province. The highest carrying rate was observed in Hainan province. Pigs at different ages displayed varying carrying rates for PCV: fattening pigs and gilts had the highest and the lowest carrying rates for PCV, respectively. In addition, the excretion rates for PCV of colostrum, semen, and nasal, anal, and vulval swabs were tested. The colostrum, anal swabs, and semen had higher excretion rates for PCV; these were followed by the vulval and nasal swabs that had excretion rates for PCV. Furthermore, a high blood virus-carrying rate was detected in moribund pigs, especially in pigs with fever and red skin. As to the virus-carrying rate in the pig organs received from clinical necropsy, the highest rate was found in placental tissue, followed by the kidneys, and the virus also was detected in lymphoid organs, liver, stomach, and intestines. The PCV2-positive samples were sequenced to reveal the molecular epidemic dynamics of PCV2. The results indicated four major branches, namely, PCV2a, PCV2b, PCV2c, and PCV2d, concerning PCV2 molecular epidemiology in China, with PCV2a, PCV2b, and PCV2d dominating. In conclusion, the results obtained in this study elucidated the molecular epidemiology, transmission, and positive blood samples of PCV and provided new ideas for developing comprehensive PCV control technologies to begin eliminating the disease caused by PCV by cleaning pig farms.

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Section: Discussionmentioning

confidence: 99%

Pathogenic ecological characteristics of PCV2 in large-scale pig farms in China affected by African swine fever in the surroundings from 2018 to 2021

Liu

Tong

et al. 2023

Front. Microbiol.

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“…In comparison, the detection limit for ddPCR was two copies/µL for PCV2 and 1copy/µL for PCV3. The detection limits of another experimental method for PCV2 and PCV3 were ten and one copies/µL (Cao, Luan, Wang, Li, & Li, 2021). Therefore, the present method provided favorable anti-interference, specificity, and repeatability and is applicable to clinical or normal diagnosis and detection, early warning, and surveillance of PCV2 and PCV3.…”

Section: Discussionmentioning

confidence: 79%

Pathogenic Ecological Characteristics of PCV2 in Large-Scale Pig Farms in China Affected by African Swine Fever from 2018-2021

Liu

Tong

et al. 2022

Preprint

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Porcine circovirus type 2 (PCV2) has been identified as the causal agent of postweaning multisystemic wasting syndrome (PMWS), an economically important multifactorial disease of the swine industry worldwide. This research designed a dual nested polymerase chain reaction (PCR) detection method to simultaneously monitor porcine circovirus type 2 (PCV2) and PCV3. The main applications for this protocol focused on technical support for its use during early warning surveillance of PCV and its application for the investigation of pathogenic ecology for large-scale pig farms in some areas of China. Two pairs of primers were designed based on the conserved regions of both PCV2 and PCV3 strain genes included in GenBank. This resulted in a highly sensitive detection method with good specificity, which was constructed by optimizing the reaction conditions and testing for specificity, sensitivity, and coincidence rate. Next, 15,130 systematic early warning and clinical samples were assessed using the developed methods. The limit of detection (LoD) of sensitivity for PCV2 and PCV3 was ten copies/mL for both viruses. There was no cross-reaction with any other porcine pathogens tested and no non-specific amplification. The coincidence and repetition rates were 100%. Through the systematic and clinical sampling, 15,130 samples from 30 large-scale pig farms in eight provinces (Hubei, Hunan, Henan, Jiangxi, Shanxi, Guangdong, Hainan, and Heilongjiang) were subjected to early warning surveillance and/or clinical diagnosis. The results revealed that the overall positive rates of PCV3 and PCV2 were 0% and 28.29%, respectively, with the lowest level [8.31% (133/1,600)] recorded in Jiangxi province. The highest carrying rate [42.36% (762/1,799)] was observed in Hainan province. Pigs at different ages displayed varying carrying rates: fattening pigs and reserve pigs had the highest and the lowest carrying rates at 65.01% (901/1,386) and 6.87% (98/1,426), respectively. In addition, the detoxification rates of colostrum, semen, and nasal, anal, and vulval swabs were tested. The colostrum, anal swabs, and semen had higher toxicity rates of 26.65%, 25.30%, and 22.71%, respectively; these were followed by the vulval and nasal swabs that had toxicity rates of 15.47% and 12.81%, respectively. Furthermore, a high blood virus-carrying rate was detected in moribund pigs, especially in pigs with fever and red skin. As to the virus-carrying rate in the pig organs received from clinical necropsy, the highest rate was found in placental tissue, followed by the kidneys, and the virus also was detected in lymphoid organs, liver, stomach, and intestines. The PCV2-positive samples were sequenced and compared to reveal the molecular epidemic dynamics of PCV2 and disclose the genetic backgrounds of these epidemic strains in China. These results provided support for tracing the sources of virus transmission in future outbreaks. The results showed that the gene sequences of the 24 PCV2 strains tested showed 95.36 to 100% homology among the detected strains and 95.46 to 99.98% among the detected strains and the reference strains. Nine strains belonged to PCV2a, seven belonged to PCV2b, two belonged to PCV2c, and six strains belonged to PCV2d.

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Establishment and Application of Dual TaqMan Fluorescence Quantitative Polymerase Chain Reaction (PCR) for the Detection of Porcine Circovirus Types 2 and 3

Cited by 2 publications

References 27 publications

Pathogenic ecological characteristics of PCV2 in large-scale pig farms in China affected by African swine fever in the surroundings from 2018 to 2021

Pathogenic ecological characteristics of PCV2 in large-scale pig farms in China affected by African swine fever in the surroundings from 2018 to 2021

Pathogenic Ecological Characteristics of PCV2 in Large-Scale Pig Farms in China Affected by African Swine Fever from 2018-2021

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